Journal: Evidence-based Complementary and Alternative Medicine : eCAM

Article Title: Total Saponins of Radix Clematis Regulate Fibroblast-Like Synoviocyte Proliferation in Rheumatoid Arthritis via the LncRNA OIP5-AS1/MiR-410-3p/Wnt7b Signaling Pathway

doi: 10.1155/2022/8393949

Figure Lengend Snippet: Effect of TSC on the expression of GSK-3 β /p-GSK-3 β protein and mRNA in FLS of AA rats. (a) The change in GSK-3 β /p-GSK-3 β protein expression was observed by immunofluorescence analysis (×200): (A) normal group, (B) model group, (C) TSC group, (D) TSC + lncRNA OIP5-AS1 siRNA group, (E) lncRNA OIP5-AS1 siRNA group, and (F) lncRNA OIP5-AS1 siRNA + NC group. (b) The change in GSK-3 β /p-GSK-3 β mRNA expression was observed by RT-qPCR. (c) The change in GSK-3 β /p-GSK-3 β protein expression was observed by Western blotting. (d) Semiquantitative analysis of GSK-3 β /p-GSK-3 β protein expression. ## P < 0.01 compared with the normal group; ∗ P < 0.05 and ∗∗ P < 0.01 compared with the model group.

Article Snippet: Diluted primary antibody: Wnt7b (Abcam, GR3241134-2 1 : 500); β -catenin (Proteintech, 00077341, 1 : 200); c-Myc (Proteintech, 00033258, 1 : 50); cyclin D1 (Abcam, GR197045-1, 1 : 50), p-GSK-3 β (Ser9) (CST, #5588, 1 : 400); SFRP4 (Wuhan Aibotek Biotechnology Co., Ltd., 9100014210, 1 : 500) was added, and samples were placed in a humid box and incubated overnight at 4°C in the dark.

Techniques: Expressing, Immunofluorescence, Quantitative RT-PCR, Western Blot

Journal: Frontiers in Cell and Developmental Biology

Article Title: β-Arrestin1 Promotes Colorectal Cancer Metastasis Through GSK-3β/β-Catenin Signaling- Mediated Epithelial-to-Mesenchymal Transition

doi: 10.3389/fcell.2021.650067

Figure Lengend Snippet: Expression of β-arrestin1 in CRC tissues and its association with the prognosis of patients with CRC. (A–C) Analysis of β-arrestin1 expression profile using data from the GSE41258 dataset. (D) Association between β-arrestin1 expression and the prognosis of patients with CRC. (E) Network representation of β-arrestin1 interactions with multiple genes, including Twist, Snail, CDH1, Vimentin, MMP2, MMP9, GSK-3β, MAPK, etc. (F–G) Immunohistochemical and quantitative analysis of β-arrestin1 protein in representative CRC and metastatic lung/liver tissues (scale bars, upper row: 200 μm, down row: 50 μm, respectively). * P < 0.05; ** P < 0.01 vs. Primary tumor.

Article Snippet: Rabbit monoclonal antibodies against human E-cadherin (cat. no. 3195; dilution 1:1,000), N-cadherin (cat. no. 13116; dilution 1:1,000), Vimentin (cat. no. 5741; dilution 1:1,000), Snail (cat. no. 3879; dilution 1:1,000), β-arrestin1 (cat. no. 12697; dilution 1:1,000), GSK-3β (cat. no. 12456; dilution 1:1,000), p-GSK-3β (cat. no. 5588; dilution 1:1,000), Axin1 (cat. no. 2087; dilution 1:1,000), CK1 (cat. no. 2655; dilution 1:1,000), β-catenin (cat. no. 8480; dilution 1:1,000), c-Myc (cat. no. 5605; dilution 1:1,000), CyclinD1 (cat. no. 2922; dilution 1:1,000) and PCNA (cat. no. 13110; dilution 1:1,000) were purchased from Cell Signaling Technology, Inc. Rabbit monoclonal antibodies against human MMP2 (cat. no. sc-13594; dilution 1:500) and MMP9 (cat. no. sc-393859; dilution 1:500) were purchased from Santa Cruz Biotechnology, Inc.

Techniques: Expressing, Immunohistochemical staining

Journal: Frontiers in Cell and Developmental Biology

Article Title: β-Arrestin1 Promotes Colorectal Cancer Metastasis Through GSK-3β/β-Catenin Signaling- Mediated Epithelial-to-Mesenchymal Transition

doi: 10.3389/fcell.2021.650067

Figure Lengend Snippet: β-arrestin1 regulates the Wnt/β-catenin signal pathway in CRC cells. (A,B) Western blotting detected the protein levels of GSK-3β, p-GSK-3β, Axin1, CK1, β-catenin, c-Myc and CyclinD1 in shRNA/β-arrestin1 and over/β-arrestin1 groups. * P < 0.05; ** P < 0.01; *** P < 0.001 vs. HCT-116-vector or LoVo-vector cells. (C,D) The protein expression of β-catenin in whole or cell extracts of different cellular compartments from HCT-116 or LoVo cells. * P < 0.05; ** P < 0.01; *** P < 0.001 vs. HCT-116-vector or LoVo-vector cells. (E,F) β-catenin inhibitor ICG001 (10 μM) was added to the over/β-arrestin1 group, and then cultured for 24 h. The associated proteins of the Wnt signaling pathway were examined via western blotting. * P < 0.05; ** P < 0.01 vs. HCT-116-vector or LoVo-vector cells. (G,H) Western blotting detected the protein expression levels of epithelial or mesenchymal markers after using ICG001. * P < 0.05; ** P < 0.01; *** P < 0.001 vs. HCT-116-vector or LoVo-vector cells. CRC, colorectal cancer.

Article Snippet: Rabbit monoclonal antibodies against human E-cadherin (cat. no. 3195; dilution 1:1,000), N-cadherin (cat. no. 13116; dilution 1:1,000), Vimentin (cat. no. 5741; dilution 1:1,000), Snail (cat. no. 3879; dilution 1:1,000), β-arrestin1 (cat. no. 12697; dilution 1:1,000), GSK-3β (cat. no. 12456; dilution 1:1,000), p-GSK-3β (cat. no. 5588; dilution 1:1,000), Axin1 (cat. no. 2087; dilution 1:1,000), CK1 (cat. no. 2655; dilution 1:1,000), β-catenin (cat. no. 8480; dilution 1:1,000), c-Myc (cat. no. 5605; dilution 1:1,000), CyclinD1 (cat. no. 2922; dilution 1:1,000) and PCNA (cat. no. 13110; dilution 1:1,000) were purchased from Cell Signaling Technology, Inc. Rabbit monoclonal antibodies against human MMP2 (cat. no. sc-13594; dilution 1:500) and MMP9 (cat. no. sc-393859; dilution 1:500) were purchased from Santa Cruz Biotechnology, Inc.

Techniques: Western Blot, shRNA, Plasmid Preparation, Expressing, Cell Culture

Journal: Scientific Reports

Article Title: Novel EGFR-targeted strategy with hybrid peptide against oesophageal squamous cell carcinoma

doi: 10.1038/srep22452

Figure Lengend Snippet: ( A ) EGFR expression levels in whole -cell lysates of HEK293 and OSCC cells (TE-5, TE-8, TE-10, TE-11, TE-5R, and TE-11R). The EGFR protein expression level was determined by Western blotting. β-Actin served as a loading control. EGFR protein was highly expressed in all OSCC cells, while its level was very low in HEK293 cells. ( B ) EGFR expressions on the cell surface of HEK293 and OSCC cells. Cells were stained with AlexaFluor 647-conjugated monoclonal anti-EGFR antibody or IgG isotype control antibody. The Fluorescence intensity determined by flow cytometry showed that EGFR was expressed on the surface of OSCC cells, whereas HEK293 cells minimally expressed it. ( C,D ) Affinity of EGFR-binding peptide for HEK293 and TE-11R cells. They were exposed to Fluorescein (FLC)-labelled EGFR-binding peptide, and the fluorescence intensity was assessed by flow cytometry. The Affinity of EGFR-binding peptide for cell membranes of TE-11R cells is increasing in time- ( C ) and dose- ( D ) dependent manners, but not in HEK293 cells. The assays were repeated three times, and data are shown as the mean ± SD. n.s.: not significant, *P < 0.05, **P < 0.01 vs. untreated (0 minutes) cells.

Article Snippet: Antibodies and the titres were as follows: AlexaFluor 647-conjugated monoclonal anti-EGFR antibody (D38B1, #5588, Cell Signaling; 1:50), rabbit IgG isotype control (Alexa Fluor 647 Conjugate) antibody (#3452, Cell Signaling; 1:50).

Techniques: Expressing, Western Blot, Staining, Fluorescence, Flow Cytometry, Binding Assay